Identification and functional characterization of new genes causing nephronophthisis: A combined genetic, evolutionary, and proteomics approach. Nephronophthisis (NPHP), an autosomal-recessive cystic kidney disease, constitutes the most frequent genetic cause of chronic renal failure in the first two decades of life. By histology, the disease is characterized by disrupted tubular basement membrane structure, renal tubular cell atrophy, interstitial fibrosis and cyst formation. In a subset of patients with NPHP there is an association with retinitis pigmentosa or liver fibrosis. We have previously identified by positional cloning two novel genes (NPHP1 and NPHP4), mutations in which cause NPHP types 1 and 4. Their gene products, nephrocystin and nephroretinin, function within matrix-epithelial cell signaling at focal adhesions and in cell-cell signaling at adherens junctions. Very recently, we have identified by positional cloning the gene (NPHP3), mutations in which cause NPHP type 3 and the mouse renal cystic phenotype pcy. In addition, using a candidate approach, we have now also identified mutations in the human inversin gene as causing NPHP type 2. Furthermore, we demonstrated that the products of all four NPHP genes are expressed in primary cilia of renal epithelial cells, and that proteins encoded by NPHP genes interact with the NPHP1 gene product nephrocystin. The NPHP1, 2 and 4 genes are conserved in evolution including the nematode C. elegans. Since all known NPHP gene products interact and are expressed in renal cilia, this proposal is aimed at the identification and characterization of new genes involved in the pathogenesis of NPHP, to elucidate the complex genetic and functional relationships of the recently and newly identified NPHP genes. Specifically, we propose to: 1. Identify and characterize further genes causing NPHP by total genome search for linkage and positional cloning; 2. Use candidate genes derived from renal cystic mouse models to identify further NPHP-causing genes; 3. Identify novel functional modules involving NPHP proteins that are conserved in evolution (C. elegans), and identify protein binding partners. Since NPHP1, 3 and 4 represent novel genes, we expect these studies to elucidate novel mechanisms of cell-matrix and cell-cell signaling and ciliary function in developing and adult kidney, and to provide new insights into disease mechanisms of renal interstitial fibrosis and cyst development as well as the function of visual function of the retina.